Gemin5 is the RNA-binding protein of the Survival of Motor Neurons (SMN) complex. The best- characterized function of the SMN complex is to assemble the spliceosomal small-nuclear RNPs (snRNPs), the major pre-mRNA splicing machinery in eukaryotes. Gemin5 binds to snRNAs and delivers then to SMN for snRNP assembly. Mutations in SMN cause Spinal Muscular Atrophy (SMA), a common and often fatal neurodegenerative disease characterized by motor-neuron degeneration and severe muscular atrophy. While SMA patients have defects in snRNP assembly and deficiency in pre-mRNA splicing, it is currently not known how this could lead to specific death of motor neurons. The specific focus of this proposal is the identification of mRNAs that bind to Gemin5. The specific RNA sequence and structural features recognized by Gemin5 are found in many different RNAs, including mRNAs. Further, Gemin5 associates with mRNA translation initiation factors. We propose here to identify specific mRNAs that bind to Gemin5 in general cell types as well as neuronal cells and SMA models. We will investigate the mechanism by which Gemin5 coordinately regulates protein expression from those mRNAs, and determine the role of SMN in that regulation. This will allow us to propose a new mechanism by which SMN and Gemin5 work together to control expression of specific proteins in SMA. The first specific aim utilizes immunoprecipitations along with microarrays and deep sequencing to undertake a global identification of the mRNAs that bind to Gemin5. We will extend this analysis to several neuronal cell types to identify Gemin5 target mRNAs that are relevant to SMA. Finally, we will use in vitro RNA-binding assays to define the affinity and sequence specificity of the Gemin5-mRNA interactions as well as the mRNA binding site on Gemin5. The second specific aim focuses on the biological role of the interaction of Gemin5 with the specific mRNAs. We will develop cell systems to explore the effect of reducing Gemin5 on protein expression from those target mRNAs. We will then investigate the mechanism by which Gemin5 regulates expression from mRNAs by examining its role in mRNA translation and localization. Finally, we will define how Gemin5 controls gene expression in SMA by using neuronal cells with reduced SMN and an SMA mouse model. PUBLIC HEALTH RELEVANCE: Gemin5 is the RNA-binding protein of the Survival of Motor Neurons (SMN) complex. Mutations in SMN cause defects in RNA metabolism and Spinal Muscular Atrophy (SMA), a common and usually fatal neurodegenerative disease. This proposal seeks to understand a new mechanism by which Gemin5 and SMN regulate gene expression in neurons, which will aid in our understanding of the pathogenesis and eventual treatment of Spinal Muscular Atrophy.